Studies are proposed to investigate the microbicidal reactions occurring in phagocytizing leukocytes. Emphasis is placed upon the cellular oxidation reactions of hypochlorous acid, which is produced by myeloperoxidase catalyzed peroxidation of chloridation. We have shown this oxidant to be highly reactive to nucleophilic biological molecules including ferredoxins, cytochromes and hemes, carotenes, nucleotides and sulfhydryl-containing enzymes and have demonstrated the occurrence of these reactions in bacterial cells (J.M. Abrich et al. (1981) Proc. Nat. Acad. Sci. USA 78, 210-214). The major objective of this proposal is to identify mechanisms of disinfection by (i) correlating loss of cellular viability, as measured by the ability to replicate, with specific changes occurring upon oxidation in various microorganisms exhibiting differing energy-generating metabolic pathways; (ii) seeking evidence for intracellular reaction sites by characterizing products from these reactions; and (iii) seeking evidence for occurrence of these reactions in stimulated neutrophils using fluorescein-labeled particles as extrinsic probes and microbial biomolecules as intrinsic probes. Particular attention will be given to changes occurring in cellular proton electrochemical potentials metabolite transport, and oxidation of nucleotides, iron-sulfur clusters and thiols, cytochromes, and carotene pigments. Major instrumentation to be used includes optical difference spectroscopy, electron paramagnetic resonance spectroscopy, and various combined chromatographic-mass spectrometric assemblies. The combined results will provide critical evaluation of the proposed role of HOC1 in phagocytic disinfection mechanisms.